Comparative enzymology of certain insect acetylesterases in relation to poisoning by organophosphorus insecticides

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Abstract
A survey was made of the enzymes hydrolyzing acetylcholine (ACh), benzoylcholine (BzCh), Acetyl-beta-methylcholine (MeCh), triacetin (TA) and o-nitrophenyl acetate (NPA) in various insects and the organ systems of the American cockroach. Substrate specificity patterns varied greatly among different insects and among organs of 1 insect. The properties of these esterases were studied by determining activities on mixed substrates and through the use of eserine, tetraethyl pyrophos-phate and choline as inhibitors in vitro. The hydrolysis of ACh, TA and NPA may be due to 1, 2 or 3 enzymes depending on the source of insect esterases. Large differences in sensitivity to the various inhibitors occurred with the various insect esterases. Both the quantity and properties of the esterases of the housefly and waxmoth shift during development. Pea aphids were shown to contain cholinesterases attacking ACh and BzCh but not MeCh. Excess substrate inhibition occurred with BzCh but not ACh. Very high activity was observed on TA and NPA and lower activity on a group of other ester substrates. The activity towards ACh could be separated from that towards NPA by solvent or salt fractionation or by the use of selective inhibitors. Enzymes very active in hydrolysing choline esters were observed in many insect parts. Noteworthy are the high activity of housefly heads and whole whiteflies for ACh, of cockroach malpighian tubules for BzCh and of cockroach ventral nerve cord for MeCh. A concentration of the tri-acetinase of the cockroach in the gastric caecae and a high activity of the blood for choline esters were also observed. The relative physiological importance of ACh esterase and NPA esterase in organophosphate poisoning of the adult cockroach was evaluated. Studies with 4 organo-phosphates, 2 substrates, and 8 organ systems showed that the only enzyme consistently inhibited to a large extent (67-85%) at the prostrate stage was the ACh esterase of the nerve cord. No evidence was obtained contrary to the hypothesis that the primary cause of organophosphate insecticide poisoning is due to blockage of the nerve acetylcholine -cholinesterase system in those insects where this system is physiologically important. Considering the specificity differences occurring in the cholinesterase part of this system among the insects and between insects and mammals, a further development of organophosphate insecticides utilizing these differences to achieve the desired degree of selective toxicity seems feasible.